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Automated System for Single Molecule Fluorescence Measurements of Surface-immobilized Biomolecules

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Metasurface Biosensors Enabling Single-Molecule Sensing of Cell-Free DNA.

Masanobu Iwanaga¹, Takashi Hironaka¹, Naoki Ikeda¹

¹National Institute for Materials Science (NIMS), 1-1 Namiki, Tsukuba 305-0044, Japan.

|June 14, 2023

Summary

Highly sensitive all-dielectric metasurface biosensors detect single cell-free DNA (cfDNA) molecules. Combining metasurface technology with reduced-cycle polymerase chain reaction (PCR) enables precise cfDNA quantification.

Keywords:

all-dielectric metasurface biosensor cfDNA fluorescence detection polymerase chain reaction single-molecule sensing

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Area of Science:

Nanotechnology and Biosensing
Molecular Diagnostics

Background:

Cell-free DNA (cfDNA) is a crucial biomarker for various physiological and pathological conditions.
Accurate detection of low-abundance cfDNA is essential for early disease diagnosis and monitoring.
Existing methods often lack the sensitivity or practicality for single-molecule cfDNA detection.

Purpose of the Study:

To develop and validate highly sensitive metasurface biosensors for single-target DNA detection.
To achieve high-precision quantification of cell-free DNA (cfDNA) using an integrated approach.
To establish a practical method for discriminating between the presence and absence of cfDNA at the single-copy level.

Main Methods:

Fabrication of all-dielectric metasurface biosensors with enhanced fluorescence (FL) properties.
Integration of metasurface biosensors with a reduced-cycle polymerase chain reaction (PCR) for nucleic acid amplification.
Single-molecule fluorescence detection and statistical analysis of FL signals.

Main Results:

Metasurface biosensors achieved single-molecule concentration detection of target DNA.
The combined approach demonstrated high statistical confidence (84% automated, 99.9% confocal microscopy) for single-molecule cfDNA detection.
A simple and practical test was developed to differentiate 1 copy/test from zero, outperforming digital PCR in this aspect.

Conclusions:

Highly fluorescence-enhancing all-dielectric metasurface biosensors offer a powerful platform for ultrasensitive cfDNA detection.
The integration with reduced-cycle PCR provides a high-precision, practical method for single-molecule cfDNA quantification.
This technology represents a significant advancement in molecular diagnostics, enabling detection at levels previously unattainable.